Many biochemical assays require labels for detection in order to convert a specific binding event into a measurable signal. Such labels may provide signals that are measurable optically, electrically, acoustically, magnetically, etc. Thus, while the binding of two molecules may be optically undetectable, if one molecule is “labeled” with an optically active species, the molecular binding event may now be detected optically.
In order to enhance the detection sensitivity of in vitro diagnostic assays, an amplification event is often performed. Labels may perform this amplification. For example, catalytic amplification may be performed by enzymes, such as horseradish peroxidase, alkaline phosphatase, etc, that are directly or indirectly bound to biological recognition molecules. Each enzyme catalyst can then produce multiple detectable molecules, resulting in an amplification of each individual biochemical recognition event.
Such enzyme labels are often utilized for signal production and amplification in enzyme immunoassays (EIAs), such as enzyme-linked immunosorbent assays (ELISAs) and enzyme-linked immunospot assays (ELISPOTs), and immunoblot assays (Western blots). Enzymes offer a limited amplification capability, may have high nonspecific binding, and can produce only a single signal, limiting their utility in multiplexed assay formats. Enzyme performance may vary significantly from batch-to-batch, over time, and with non-optimal storage conditions. Furthermore, enzyme conjugates, specifically enzyme-antibody conjugates, are difficult to manufacture, adding the requirement of at least one additional detection step in some assay formats. Existing non-enzymatic amplification strategies suffer from low amplification levels, high non-specific and off-target binding, and/or the use of non-standard equipment, limiting their performance and utility.